The mechanism of action of beef heart mitochondrial ATPase will be studied. A direct examination will be carried out of the properties of the two hydrolytic sites on each molecule of the enzyme. An effort will be made to determine if a third hydrolytic site, predicted from genetic and other evidence, is present on the protein. A study of the fate of 18O-labelled adenine nucleotide and analogs during hydrolysis by F1 will be undertaken in an effort to confirm current thinking in this laboratory that site-site cooperativity in the action of F1 occurs in the bond breaking step of hydrolysis rather than the product release step.